bims-prodis 2019-03-03 papers

bims-prodis

Biomed News

on Proteomics in disease

Issue of 2019‒03‒03
twenty-five papers selected by
Nancy Gough
Bioserendipity

Urinary Proteomics Profiles Are Useful for Detection of Cancer Biomarkers and Changes Induced by Therapeutic Procedures.
Proteomic maps of human gastrointestinal stromal tumor subgroups.
Plasma proteome profiling discovers novel proteins associated with non-alcoholic fatty liver disease.
Identification of SAA and ACTB as potential biomarker of patients with severe HFMD using iTRAQ quantitative proteomics.
Comparative Proteome-Wide Analysis of Bone Marrow Microenvironment of β-Thalassemia/Hemoglobin E.
Differential urinary proteins to diagnose coronary heart disease based on iTRAQ quantitative proteomics.
Protein profiling of osteosarcoma tissue and soft callus unveils activation of the unfolded protein response pathway.
Differential epithelial and stromal protein profiles in cone and non-cone regions of keratoconus corneas.
Mediators of Corneal Haze Following Implantation of Presbyopic Corneal Inlays.
Dynamic protein changes in the perihaemorrhagic zone of Surgically Treated Intracerebral Haemorrhage Patients.
Locally different proteome in aortas from patients with stenotic tricuspid and bicuspid aortic valves†.
Proteomics identifies new therapeutic targets of early-stage hepatocellular carcinoma.
Proteomics Analysis of FLT3-ITD Mutation in Acute Myeloid Leukemia Using Deep Learning Neural Network.
GLAST (GLutamate and ASpartate Transporter) in human prefrontal cortex; interactome in healthy brains and the expression of GLAST in brains of chronic alcoholics.
Longitudinal effects of aging on plasma proteins levels in older adults - associations with kidney function and hemoglobin levels.
Oxidative Stress and Bronchopulmonary Dysplasia: Evidences From Microbiomics, Metabolomics, and Proteomics.
Multiplexed Immunohistochemistry for Molecular and Immune Profiling in Lung Cancer-Just About Ready for Prime-Time?
Highly multiplexed profiling of cell surface proteins on single circulating tumor cells based on antibody and cellular barcoding.
Molecular profile of ultrastructure changes of the ligamentum flavum related to lumbar spinal canal stenosis.
Evaluating Metagenomic Prediction of the Metaproteome in a 4.5-Year Study of a Patient with Crohn's Disease.
Proteomics reveals profound metabolic changes in the alcohol use disorder brain.
Ibudilast sensitizes glioblastoma to temozolomide by targeting Macrophage Migration Inhibitory Factor (MIF).
Re-assessment of the high-grade serous ovarian cancer phosphoproteome - Identification of kinase candidates in TCGA tumors for future clinical intervention.
AF1q gene polymorphism proteomic markers in herpes zoster-infected leukemia patients.
Neutrophil exosomes enhance the skin autoinflammation in generalized pustular psoriasis via activating keratinocytes.

Molecules. 2019 Feb 22. pii: E794. [Epub ahead of print]24(4):

Urinary Proteomics Profiles Are Useful for Detection of Cancer Biomarkers and Changes Induced by Therapeutic Procedures.

Emanuele Ferrari, Andrea Wittig, Fabrizio Basilico, Rossana Rossi, Antonella De Palma, Dario Di Silvestre, Wolfgang A G Sauerwein, Pier Luigi Mauri.

  Boron neutron capture therapy (BNCT) is a binary cancer treatment modality where two different agents (10B and thermal neutrons) have to be present to produce an effect. A dedicated trial design is necessary for early clinical trials. The concentration of 10B in tissues is an accepted surrogate to predict BNCT effects on tissues. Tissue, blood, and urines were sampled after infusion of two different boron carriers, namely BSH and BPA in the frame of the European Organisation for Research and Treatment of Cancer (EORTC) trial 11001. In this study, urine samples were used to identify protein profiles prior and after drug infusion during surgery. Here, an approach that is based on the mass spectrometry (MS)-based proteomic analysis of urine samples from head and neck squamous cell carcinoma (HNSCC) and thyroid cancer patients is presented. This method allowed the identification of several inflammation- and cancer-related proteins, which could serve as tumor biomarkers. In addition, changes in the urinary proteome during and after therapeutic interventions were detected. In particular, a reduction of three proteins that were involved in inflammation has been observed: Galectin-3 Binding Protein, CD44, and osteopontin. The present work represents a proof of principle to follow proteasome changes during complex treatments based on urine samples.

Keywords:  BNCT; LC-MS; MudPIT; boron; proteomics; squamous cell cancer of head and neck; thyroid cancer; urine

DOI:  https://doi.org/10.3390/molecules24040794
Mol Cell Proteomics. 2019 Feb 25. pii: mcp.RA119.001361. [Epub ahead of print]

Proteomic maps of human gastrointestinal stromal tumor subgroups.

Yu Liu, Zhigui Li, Zhiqiang Xu, Xiuxiu Jin, Yanqiu Gong, Xuyang Xia, Yuqin Yao, Zhaofen Xu, Yong Zhou, Heng Xu, Shuangqing Li, Yong Peng, Xiaoting Wu, Lunzhi Dai.

  Gastrointestinal stromal tumor (GIST) is a common sarcoma of gastrointestinal tract (GIT) with high metastatic and recurrence rates, but the proteomic features are still less understood. Here we performed systematic quantitative proteome profiling of GIST from 13 patients classified into very low/low, intermediate and high risk subgroups. An extended cohort of GIST (n = 131) was used for immunohistochemical validation of proteins of interest. In total, 9177 proteins were quantified, covering 55.9% of the GIT transcriptome from The Human Protein Altas. Out of the 9177 quantified proteins, 4930 proteins were observed in all 13 cases with 517 upregulated and 187 downregulated proteins in tumorous tissues independent of risk stage. Pathway analysis showed that the downregulated proteins were mostly enriched in metabolic pathway, while the upregulated proteins mainly belonged to spliceosome pathway. In addition, 131 proteins showed differentially expressed patterns among GIST subgroups with statistical significance. The 13 GIST cases were classified into 3 subgroups perfectly based on the expression of these proteins. The intensive comparison of molecular phenotypes and possible functions of quantified oncoproteins, tumor suppressors, phosphatases and kinases between GIST subgroups was carried out. Immunohistochemical analysis of the phosphatase PTPN1 (n = 117) revealed that the GIST patients with high PTPN1 expression had low chances of developing metastasis. Collectively, this work provides valuable information for understanding the inherent biology and evolution of GIST.

Keywords:  Clinical proteomics; Diagnostic; GIST; Gastrointestinal disease; High Throughput Screening; Immunohistochemistry; Mass Spectrometry; PPP2CB; PTPN1; Protein Phosphatases*; Quantitative proteomics; risk classification

DOI:  https://doi.org/10.1074/mcp.RA119.001361
Mol Syst Biol. 2019 Mar 01. 15(3): e8793

Plasma proteome profiling discovers novel proteins associated with non-alcoholic fatty liver disease.

Lili Niu, Philipp E Geyer, Nicolai J Wewer Albrechtsen, Lise L Gluud, Alberto Santos, Sophia Doll, Peter V Treit, Jens J Holst, Filip K Knop, Tina Vilsbøll, Anders Junker, Stephan Sachs, Kerstin Stemmer, Timo D Müller, Matthias H Tschöp, Susanna M Hofmann, Matthias Mann.

  Non-alcoholic fatty liver disease (NAFLD) affects 25% of the population and can progress to cirrhosis with limited treatment options. As the liver secretes most of the blood plasma proteins, liver disease may affect the plasma proteome. Plasma proteome profiling of 48 patients with and without cirrhosis or NAFLD revealed six statistically significantly changing proteins (ALDOB, APOM, LGALS3BP, PIGR, VTN, and AFM), two of which are already linked to liver disease. Polymeric immunoglobulin receptor (PIGR) was significantly elevated in both cohorts by 170% in NAFLD and 298% in cirrhosis and was further validated in mouse models. Furthermore, a global correlation map of clinical and proteomic data strongly associated DPP4, ANPEP, TGFBI, PIGR, and APOE with NAFLD and cirrhosis. The prominent diabetic drug target DPP4 is an aminopeptidase like ANPEP, ENPEP, and LAP3, all of which are up-regulated in the human or mouse data. Furthermore, ANPEP and TGFBI have potential roles in extracellular matrix remodeling in fibrosis. Thus, plasma proteome profiling can identify potential biomarkers and drug targets in liver disease.

Keywords:   NAFLD ; NASH ; biomarker discovery; mass spectrometry; plasma proteome profiling

DOI:  https://doi.org/10.15252/msb.20188793
Clin Biochem. 2019 Feb 25. pii: S0009-9120(18)30769-0. [Epub ahead of print]

Identification of SAA and ACTB as potential biomarker of patients with severe HFMD using iTRAQ quantitative proteomics.

Fangye Zhou, Xianqian Chen, Guoxian Chen, Junhua Yan, Yupeng Xiao.

  Hand, foot and mouth disease (HFMD) is an infectious disease caused by a variety of enterovirus infections, and the most common types of virus infections are the newenterovirus71 (EV71) and coxsackievirus A group 16 (CoxA16). A small fraction of HFMD will cause further severe HFMD. A rapid and accurate diagnosis biomarker of severe HFMD is important for the timely treatment. In the study, we conducted a clinical biomarker discovery study using iTRAQ combined with MS. Serum proteome alterations in severe HFMD group (n = 32) and health control group (n = 32) were analyzed. 47 proteins were upregulated (fold change > 1.5) between the severe HFMD group and HC group. The identified proteins were classified into different groups according to the molecular function, biology processes, cellular component. During the up-regulated proteins, serum amyloid A (SAA) and human β-actin (ACTB), were confirmed in the serum of the severe HFMD and HC by ELISA assay. SAA and ACTB levels were significantly higher in the sever HFMD patients (P < .01), consistent with iTRAQ-LC-MS/MS analysis. In summary, Our results showed that SAA and human β-actin (ACTB) may be served as a potential biomarker of the clinical diagnosis of severe HFMD.

Keywords:  Biomarker; Foot and mouth disease; Hand; Human β-actin; Serum amyloid A

DOI:  https://doi.org/10.1016/j.clinbiochem.2019.02.011
Proteomes. 2019 Feb 23. pii: E8. [Epub ahead of print]7(1):

Comparative Proteome-Wide Analysis of Bone Marrow Microenvironment of β-Thalassemia/Hemoglobin E.

Saranyoo Ponnikorn, Rungrawee Mongkolrob, Suwit Klongthalay, Sittiruk Roytrakul, Kitima Srisanga, Sumalee Tungpradabkul, Suradej Hongeng.

  β-thalassemia/Hb E is a global health issue, which is characterized by a range of clinical symptoms from a mild and asymptomatic anemia to severe disorders that require transfusions from infancy. Pathological mechanisms of the disease involve the excess of unmatched alpha globin and iron overload, leading to ineffective erythropoiesis and ultimately to the premature death of erythroid precursors in bone marrow (BM) and peripheral organs. However, it is unclear as to how BM microenvironment factors contribute to the defective erythropoiesis in β-thalassemia/Hb E patients. Here, we employed mass spectrometry-based comparative proteomics to analyze BM plasma that was collected from six β-thalassemia/Hb E patients and four healthy donors. We identified that the differentially expressed proteins are enriched in secretory or exosome-associated proteins, many of which have putative functions in the oxidative stress response. Using Western blot assay, we confirmed that atypical lipoprotein, Apolipoprotein D (APOD), belonging to the Lipocalin transporter superfamily, was significantly decreased in BM plasma of the tested pediatric β-thalassemia/Hb E patients. Our results highlight that the disease condition of ineffective erythropoiesis and oxidative stress found in BM microenvironment of β-thalassemia/Hb E patients is associated with the impaired expression of APOD protein.

Keywords:  Apolipoprotein D; bone marrow; ineffective erythropoiesis; oxidative stress; proteomics; β-thalassemia/Hb E

DOI:  https://doi.org/10.3390/proteomes7010008
Anal Bioanal Chem. 2019 Feb 26.

Differential urinary proteins to diagnose coronary heart disease based on iTRAQ quantitative proteomics.

Haidan Sun, Danqi Wang, Dongfang Liu, Zhengguang Guo, Chen Shao, Wei Sun, Yong Zeng.

  Coronary artery disease (CAD) is a manifestation of systemic atherosclerotic disease. It is assessed by intervention or traditional scoring risk factors. Diagnosis is limited by inaccurate and invasive methods. Developing noninvasive methods to screen for the risk of CAD is a major challenge. We aimed to identify urinary proteins associated with CAD. We utilized iTRAQ labeling followed by 2D LC-MS/MS to compare the urinary proteome of CAD patients to healthy cohorts. The multiple reaction monitoring (MRM) was used to verify the differential proteins. ROC analysis based on MRM data was used to evaluate the diagnostic application. A total of 876 proteins were quantified, and 100 differential proteins were found. Functional analysis revealed that the differential proteins were mainly associated with Liver X Receptor/Retinoid X Receptor (LXR/RXR) pathway activation, atherosclerosis signaling, production of nitric oxide and reactive oxygen species, and the top upstream regulator of the differential proteins by IPA analysis indicated to the APOE. Nineteen differential proteins were verified by MRM analysis. ROC based on MRM data revealed that the combination of two proteins (APOD and TFF1) could diagnose CAD with 85% sensitivity and 99% specificity (AUC 0.95). The urinary proteome might reflect the pathophysiological changes in CAD and be used for the clinical study of CAD.

Keywords:  Atherosclerosis; MRM; Proteomics; Urine; iTRAQ

DOI:  https://doi.org/10.1007/s00216-019-01668-7
Int J Oncol. 2019 Feb 28.

Protein profiling of osteosarcoma tissue and soft callus unveils activation of the unfolded protein response pathway.

Parunya Chaiyawat, Patsadakorn Sungngam, Pimpisa Teeyakasem, Nutnicha Sirikaew, Jeerawan Klangjorhor, Jongkolnee Settakorn, Penchatr Diskul-Na-Ayudthaya, Daranee Chokchaichamnankit, Chantragan Srisomsap, Jisnuson Svasti, Dumnoensun Pruksakorn.

Oncogenic drivers of osteosarcoma remain controversial due to the complexity of the genomic background of the disease. There are limited novel therapeutic options, and the survival rate of patients with osteosarcoma has not improved in decades. Genomic instability leads to complexity in various pathways, which is potentially revealed at the protein level. Therefore, the present study aimed to identify the mechanisms involved in the oncogenesis of osteosarcoma using proteomics and bioinformatics tools. As clinical specimens from patients are the most relevant disease‑related source, expression patterns of proteins in osteosarcoma tissues were compared with soft tissue callus from donors containing high numbers of osteoblastic cells. Two‑dimensional electrophoresis and liquid chromatography‑tandem mass spectrometry (LC‑MS/MS) successfully identified 33 differentially expressed proteins in the osteosarcoma tissues compared with the soft tissue callus. Among these proteins, 29 proteins were significantly upregulated in osteosarcoma. A functionally grouped network of the overexpressed proteins, that was created using the ClueGo and CluePedia applications, demonstrated that the unfolded protein response (UPR) pathway was activated mainly through the activating transcription factor 6 arm in osteosarcoma. The results of proteomics analysis were confirmed by elevated expression of UPR‑related chaperone proteins, including 78 kDa glucose‑related protein (GRP78), endoplasmin, calreticulin and prelamin‑A/C, in the patient‑derived primary cells and osteosarcoma cell lines. Furthermore, the expression of GRP78, a master regulator of the UPR, was enhanced in the osteosarcoma tissues of patients that were resistant to double regimen of doxorubicin and a platinum‑based drug. The findings of the present study suggest that targeting the UPR pathway may be promising for the treatment of osteosarcoma.

DOI: https://doi.org/10.3892/ijo.2019.4737
Sci Rep. 2019 Feb 27. 9(1): 2965

Differential epithelial and stromal protein profiles in cone and non-cone regions of keratoconus corneas.

Gary Hin-Fai Yam, Matthias Fuest, Lei Zhou, Yu-Chi Liu, Lu Deng, Anita Sook-Yee Chan, Hon Shing Ong, Wei-Boon Khor, Marcus Ang, Jodhbir S Mehta.

Keratoconus (KC) is an ectatic corneal disease characterized by progressive thinning and irregular astigmatism, and a leading indication for corneal transplantation. KC-associated changes have been demonstrated for the entire cornea, but the pathological thinning and mechanical weakening is usually localized. We performed quantitative proteomics using Sequential Windowed Acquisition of All Theoretical Fragment Ion Mass Spectrometry (SWATH-MS) to analyze epithelial and stromal changes between the topographically-abnormal cone and topographically-normal non-cone regions of advanced KC corneas, compared to age-matched normal corneas. Expression of 20 epithelial and 14 stromal proteins was significantly altered (≥2 or ≤0.5-fold) between cone and non-cone in all 4 KC samples. Ingenuity pathway analysis illustrated developmental and metabolic disorders for the altered epithelial proteome with mitochondrion as the significant gene ontology (GO) term. The differential stromal proteome was related to cellular assembly, tissue organization and connective tissue disorders with endoplasmic reticulum protein folding as the significant GO term. Validation of selected protein expression was performed on archived KC, non-KC and normal corneal specimens by immunohistochemistry. This is the first time to show that KC-associated proteome changes were not limited to the topographically-thinner and mechanically-weakened cone but also non-cone region with normal topography, indicating a peripheral involvement in KC development.

DOI: https://doi.org/10.1038/s41598-019-39182-6
Invest Ophthalmol Vis Sci. 2019 Mar 01. 60(4): 868-876

Mediators of Corneal Haze Following Implantation of Presbyopic Corneal Inlays.

Beau J Fenner, Yu-Chi Liu, Siew Kwan Koh, Yan Gao, Lu Deng, Roger W Beuerman, Lei Zhou, Julian T S Theng, Jodhbir S Mehta.

Purpose: To identify protein mediators of corneal haze following presbyopic corneal inlay surgery.Methods: Tears were collected from eyes with corneal haze following surgery with a shape-changing corneal inlay. Samples were subjected to quantitative proteomic analysis using iTRAQ and proteins significantly increased or decreased (1.3-fold or more) in haze eyes relative to fellow eyes were identified. Expression ratios were compared to postoperative eyes without corneal haze to identify proteins selectively increased or decreased in corneal haze eyes.
Results: Inlay-associated haze occurred in 35% of eyes (6 of 17). Of 1443 unique tear proteins identified, eight proteins were selectively reduced in tears from postoperative haze eyes and one protein selectively increased. Proteins reduced in haze eyes included complement 4a (level relative to nonhaze eyes 0.464, P = 0.037), complement factor H (0.589, P = 0.048), immunoglobulin kappa variable 2-29 (0.128, P = 0.006), immunoglobulin kappa variable 2D-28 (0.612, P = 0.025), immunoglobulin lambda variable 7-46 (0.482, P = 0.007), S100 calcium binding protein A4 (0.614, P = 0.048), Shootin-1 (0.614, P = 0.048), and tissue inhibitor of metalloproteinase-1 (0.736, P = 0.023). The Xaa-Pro aminopeptidase 1 was increased in haze eyes relative to nonhaze eyes (1.517, P = 0.023).
Conclusions: Corneal haze following corneal inlay surgery is associated with reduction in levels of known inflammatory and immune mediators. These findings represent a starting point for elucidation of pathways involved in corneal haze following synthetic inlay implantation and may enable development of targeted therapies that modulate the haze response.

DOI: https://doi.org/10.1167/iovs.18-25761
Sci Rep. 2019 Feb 28. 9(1): 3181

Dynamic protein changes in the perihaemorrhagic zone of Surgically Treated Intracerebral Haemorrhage Patients.

Lovisa Tobieson, Bijar Ghafouri, Peter Zsigmond, Sandro Rossitti, Jan Hillman, Niklas Marklund.

The secondary injury cascades exacerbating the initial brain injury following intracerebral haemorrhage (ICH) are incompletely understood. We used dual microdialysis (MD) catheters placed in the perihaemorrhagic zone (PHZ) and in seemingly normal cortex (SNX) at time of surgical ICH evacuation in ten patients (range 26-70 years). Routine interstitial MD markers (including glucose and the lactate/pyruvate ratio) were analysed and remaining microdialysate was analysed by two-dimensional gel electrophoresis (2-DE) and nano-liquid chromatography tandem mass spectrometry (nLC-MS/MS). Two time intervals were analysed; median 2-10 hours post-surgery (time A) and median 68-76 hours post-ICH onset (time B). Using 2-DE, we quantified 232 ± 31 different protein spots. Two proteins differed between the MD catheters at time A, and 12 proteins at time B (p < 0.05). Thirteen proteins were significantly altered between time A and time B in the SNX and seven proteins in the PHZ, respectively. Using nLC-MS/MS ca 800 proteins were identified out of which 76 were present in all samples. At time A one protein was upregulated and two downregulated, and at time B, seven proteins were upregulated, and four downregulated in the PHZ compared to the SNX. Microdialysis-based proteomics is feasible for study of secondary injury mechanisms and discovery of biomarkers after ICH.

DOI: https://doi.org/10.1038/s41598-019-39499-2
Eur J Cardiothorac Surg. 2019 Feb 25. pii: ezz032. [Epub ahead of print]

Locally different proteome in aortas from patients with stenotic tricuspid and bicuspid aortic valves†.

Amalia Forte, Xiaoke Yin, Marika Fava, Ciro Bancone, Marilena Cipollaro, Marisa De Feo, Manuel Mayr, Marjan Jahangiri, Alessandro Della Corte.

  OBJECTIVES: We aimed to compare the intracellular proteome of ascending aortas from patients with stenotic bicuspid (BAV) and tricuspid aortic valves (TAV) to identify BAV-specific pathogenetic mechanisms of aortopathy and to verify the previously reported asymmetric expression of BAV aortopathy [concentrated at the convexity (CVX)] in its 'ascending phenotype' form.METHODS: Samples were collected from the CVX and concavity sides of non-aneurysmal ascending aortas in 26 TAV and 26 BAV patients undergoing stenotic aortic valve replacement. Aortic lysates were subjected to cellular protein enrichment by subfractionation, and to proteome comparison by 2-dimensional fluorescence difference in-gel electrophoresis. Differentially regulated protein spots were identified by liquid chromatography-tandem mass spectrometry and analysed in silico. Selected results were verified by immunofluorescence and reverse transcription-polymerase chain reaction.
RESULTS: In BAV samples, 52 protein spots were differentially regulated versus TAV samples at the CVX and 10 spots at the concavity: liquid chromatography-tandem mass spectrometry identified 35 and 10 differentially regulated proteins, respectively. Charge trains of individual proteins (e.g. annexins) suggested the presence of post-translational modifications possibly modulating their activity. At the CVX, 37 of the 52 different protein spots showed decreased expression in BAV versus TAV. The affected biological pathways included those involved in smooth muscle cell contractile phenotype, metabolism and cell stress.
CONCLUSIONS: The observed differential proteomics profiles may have a significant impact on the pathogenesis of the aortopathy, pointing the way for further studies. At a preaneurysmal stage, an aorta with BAV shows more protein expression changes and potentially more post-translational modifications at the CVX of the ascending aorta than at the concavity, compared to that of TAV.

Keywords:  2-dimensional fluorescence difference in-gel electrophoresis; Aortopathy; Bicuspid aortic valve; Liquid chromatography–tandem mass spectrometry; Proteome subfractionation

DOI:  https://doi.org/10.1093/ejcts/ezz032
Nature. 2019 Feb 27.

Proteomics identifies new therapeutic targets of early-stage hepatocellular carcinoma.

Ying Jiang, Aihua Sun, Yang Zhao, Wantao Ying, Huichuan Sun, Xinrong Yang, Baocai Xing, Wei Sun, Liangliang Ren, Bo Hu, Chaoying Li, Li Zhang, Guangrong Qin, Menghuan Zhang, Ning Chen, Manli Zhang, Yin Huang, Jinan Zhou, Yan Zhao, Mingwei Liu, Xiaodong Zhu, Yang Qiu, Yanjun Sun, Cheng Huang, Meng Yan, Mingchao Wang, Wei Liu, Fang Tian, Huali Xu, Jian Zhou, Zhenyu Wu, Tieliu Shi, Weimin Zhu, Jun Qin, Lu Xie, Jia Fan, Xiaohong Qian, Fuchu He, .

Hepatocellular carcinoma is the third leading cause of deaths from cancer worldwide. Infection with the hepatitis B virus is one of the leading risk factors for developing hepatocellular carcinoma, particularly in East Asia1. Although surgical treatment may be effective in the early stages, the five-year overall rate of survival after developing this cancer is only 50-70%2. Here, using proteomic and phospho-proteomic profiling, we characterize 110 paired tumour and non-tumour tissues of clinical early-stage hepatocellular carcinoma related to hepatitis B virus infection. Our quantitative proteomic data highlight heterogeneity in early-stage hepatocellular carcinoma: we used this to stratify the cohort into the subtypes S-I, S-II and S-III, each of which has a different clinical outcome. S-III, which is characterized by disrupted cholesterol homeostasis, is associated with the lowest overall rate of survival and the greatest risk of a poor prognosis after first-line surgery. The knockdown of sterol O-acyltransferase 1 (SOAT1)-high expression of which is a signature specific to the S-III subtype-alters the distribution of cellular cholesterol, and effectively suppresses the proliferation and migration of hepatocellular carcinoma. Finally, on the basis of a patient-derived tumour xenograft mouse model of hepatocellular carcinoma, we found that treatment with avasimibe, an inhibitor of SOAT1, markedly reduced the size of tumours that had high levels of SOAT1 expression. The proteomic stratification of early-stage hepatocellular carcinoma presented in this study provides insight into the tumour biology of this cancer, and suggests opportunities for personalized therapies that target it.

DOI: https://doi.org/10.1038/s41586-019-0987-8
Ann Clin Lab Sci. 2019 Jan;49(1): 119-126

Proteomics Analysis of FLT3-ITD Mutation in Acute Myeloid Leukemia Using Deep Learning Neural Network.

Christine A Liang, Lei Chen, Amer Wahed, Andy N D Nguyen.

Deep Learning can significantly benefit cancer proteomics and genomics. In this study, we attempted to determine a set of critical proteins that were associated with the FLT3-ITD mutation in newly-diagnosed acute myeloid leukemia patients. A Deep Learning network consisting of autoencoders formed a hierarchical model from which high-level features were extracted without labeled training data. Dimensional reduction reduced the number of critical proteins from 231 to 20. Deep Learning found an excellent correlation between FLT3-ITD mutation with the levels of these 20 critical proteins (accuracy 97%, sensitivity 90%, and specificity 100%). Our Deep Learning network could hone in on 20 proteins with the strongest association with FLT3-ITD. The results of this study allow for a novel approach to determine critical protein pathways in the FLT3-ITD mutation, and provide proof-of-concept for an accurate approach to model big data in cancer proteomics and genomics.

Keywords: AML; Deep Learning; FLT3-ITD; Neural Network; Proteomics
Neurochem Int. 2019 Feb 25. pii: S0197-0186(18)30613-2. [Epub ahead of print]

GLAST (GLutamate and ASpartate Transporter) in human prefrontal cortex; interactome in healthy brains and the expression of GLAST in brains of chronic alcoholics.

Mohammed Abul Kashem, Nilufa Sultana, David V Pow, Vladimir J Balcar.

  We have analysed post-mortem samples of prefrontal cortex from control and alcoholic human brains by the technique of Western blotting to estimate and compare the expressions of glutamate transporter GLAST (Excitatory Amino Acid Transporter One; EAAT1). Furthermore, using the non-alcoholic prefrontal cortex and custom-made GLAST (EAAT1) antibody we determined GLAST (EAAT1) "interactome" i.e. the set of proteins selectively bound by GLAST (EAAT1). We found that GLAST (EAAT1) was significantly more abundant (about 1.6-fold) in the cortical tissue from alcoholic brains compared to that from non-alcoholic controls. The greatest increase in the level of GLAST (EAAT1) was found in plasma membrane fraction (2.2-fold). Additionally, using the prefrontal cortical tissue from control brains, we identified 38 proteins specifically interacting with GLAST (EAAT1). These can be classified as contributing to the cell structure (6 proteins; 16%), energy and general metabolism (18 proteins; 47%), neurotransmitter metabolism (three proteins; 8%), signalling (6 proteins: 16%), neurotransmitter storage/release at synapses (three proteins; 8%) and calcium buffering (two proteins; 5%). We discuss possible consequences of the increased expression of GLAST (EAAT1) in alcoholic brain tissue and whether or how this could disturb the function of the proteins potentially interacting with GLAST (EAAT1) in vivo. The data represent an extension of our previous proteomic and metabolomic studies of human alcoholism revealing another aspect of the complexity of changes imposed on brain by chronic long-term consumption of ethanol.

Keywords:  Alcohol; Glutamate transport; Human brain; Interactome; Prefrontal cortex; Proteomics

DOI:  https://doi.org/10.1016/j.neuint.2019.02.009
PLoS One. 2019 ;14(2): e0212060

Longitudinal effects of aging on plasma proteins levels in older adults - associations with kidney function and hemoglobin levels.

Lars Lind, Johan Sundström, Anders Larsson, Erik Lampa, Johan Ärnlöv, Erik Ingelsson.

BACKGROUND: A targeted proteomics chip has been shown to be useful to discover novel associations of proteins with cardiovascular disease. We investigated how these proteins change with aging, and whether this change is related to a decline in kidney function, or to a change in hemoglobin levels.MATERIAL AND METHODS: In the Prospective Investigation of the Vasculature in Uppsala Seniors (PIVUS) study, including 1,016 participants from the general population aged 70 at baseline, 84 proteins were measured at ages 70, 75, 80. At these occasions, glomerular filtration rate (eGFR) was estimated and the hemoglobin levels were measured.
RESULTS: Sixty-one of the 84 evaluated proteins changed significantly during the 10-year follow-up (multiple testing-adjusted alpha = 0.00059), most showing an increase. The change in eGFR was inversely related to changes of protein levels for the vast majority of proteins (74%). The change in hemoglobin was significantly related to the change in 40% of the evaluated proteins, with no obvious preference of the direction of these relationships.
CONCLUSION: The majority of evaluated proteins increased with aging in adults. Therefore, normal ranges for proteins might be given in age-strata. The increase in protein levels was associated with the degree of reduction in eGFR for the majority of proteins, while no clear pattern was seen for the relationships between the proteins and the change in hemoglobin levels. Studies on changes in urinary proteins are warranted to understand the association between the reduction in eGFR and increase in plasma protein levels.

DOI: https://doi.org/10.1371/journal.pone.0212060
Front Pediatr. 2019 ;7 30

Oxidative Stress and Bronchopulmonary Dysplasia: Evidences From Microbiomics, Metabolomics, and Proteomics.

Letizia Capasso, Giovanni Vento, Cristina Loddo, Chiara Tirone, Federica Iavarone, Francesco Raimondi, Carlo Dani, Vassilios Fanos.

  Bronchopulmonary dysplasia is a major issue affecting morbidity and mortality of surviving premature babies. Preterm newborns are particularly susceptible to oxidative stress and infants with bronchopulmonary dysplasia have a typical oxidation pattern in the early stages of this disease, suggesting the important role of oxidative stress in its pathogenesis. Bronchopulmonary dysplasia is a complex disease where knowledge advances as new investigative tools become available. The explosion of the "omics" disciplines has recently affected BPD research. This review focuses on the new evidence coming from microbiomics, metabolomics and proteomics in relation to oxidative stress and pathogenesis of bronchopulmonary dysplasia. Since the pathogenesis is not yet completely understood, information gained in this regard would be important for planning an efficacious prevention and treatment strategy for the future.

Keywords:  bronchopulmonary dysplasia; metabolomics; microbiomics; newborn; oxidative stress; preterm; proteomics

DOI:  https://doi.org/10.3389/fped.2019.00030
Cancers (Basel). 2019 Feb 27. pii: E283. [Epub ahead of print]11(3):

Multiplexed Immunohistochemistry for Molecular and Immune Profiling in Lung Cancer-Just About Ready for Prime-Time?

Paul Hofman, Cécile Badoual, Fiona Henderson, Léa Berland, Marame Hamila, Elodie Long-Mira, Sandra Lassalle, Hélène Roussel, Véronique Hofman, Eric Tartour, Marius Ilié.

  As targeted molecular therapies and immuno-oncology have become pivotal in the management of patients with lung cancer, the essential requirement for high throughput analyses and clinical validation of biomarkers has become even more intense, with response rates maintained in the 20%⁻30% range. Moreover, the list of treatment alternatives, including combination therapies, is rapidly evolving. The molecular profiling and specific tumor-associated immune contexture may be predictive of response or resistance to these therapeutic strategies. Multiplexed immunohistochemistry is an effective and proficient approach to simultaneously identify specific proteins or molecular abnormalities, to determine the spatial distribution and activation state of immune cells, as well as the presence of immunoactive molecular expression. This method is highly advantageous for investigating immune evasion mechanisms and discovering potential biomarkers to assess mechanisms of action and to predict response to a given treatment. This review provides views on the current technological status and evidence for clinical applications of multiplexing and how it could be applied to optimize clinical management of patients with lung cancer.

Keywords:  brightfield; chromogenic; digital; fluorescence; immune profiling; immune-oncology; lung cancer; molecular; multiplexed

DOI:  https://doi.org/10.3390/cancers11030283
Anal Bioanal Chem. 2019 Feb 28.

Highly multiplexed profiling of cell surface proteins on single circulating tumor cells based on antibody and cellular barcoding.

Chunying Wang, Liu Yang, Zhuo Wang, Jianjun He, Qihui Shi.

  Circulating tumor cells (CTCs) are extraordinarily rare in blood samples and represent a real-time "liquid biopsy" of tumors. Although genetic and transcriptional sequencing of single CTCs has been reported, these methods fail to provide phenotypic and functional information of CTCs such as protein levels of surface proteins. Studies of single-cell proteomic assays of CTCs have been rare because of a lack of single-cell proteomic methods to handle and analyze rare cells in a high background of non-target cells with high sensitivity, throughput, and multiplexing capacity. Here, we develop a microchip-assisted single-cell proteomic method for profiling surface proteins of CTCs based on antibody and cellular DNA barcoding strategy. We combine DNA-encoded antibody tags and cell indexes to profile 15 proteins in ~ 100 single rare cells simultaneously, and use high-throughput sequencing as the readout to generate surface protein profiles of CTCs according to their cell indexes and antibody-derived protein barcodes. A 6400-well microchip and the automated puncher are used to rapidly retrieve single CTCs from enriched CTC population with minimal cell loss (~ 10%). This technological platform integrates reliable isolation and proteomic analysis of single CTCs and can be extendable to ~ 100 proteins in hundreds of rare cells with single-cell precision.

Keywords:  Circulating tumor cells; DNA barcoding; High-throughput sequencing; Microchip; Single-cell proteomic analysis

DOI:  https://doi.org/10.1007/s00216-019-01666-9
J Cell Biochem. 2019 Mar 01.

Molecular profile of ultrastructure changes of the ligamentum flavum related to lumbar spinal canal stenosis.

Eliezer Sidon, Shai S Shemesh, Lisa Mor-Yossef Moldovan, Yarden Wiesenfeld, Nissim Ohana, Dafna Benayahu.

  Lumbar spinal canal stenosis (LSCS) is a degenerative disease observed by hypertrophy of the ligamentum flavum (LF) that cause compression of the lumbar neural content. Diabetes mellitus (DM) is a risk factor for the disease and we have shown previously that DM increases the fibrosis and elastic fiber loss in patients with LSCS. The purpose of this study was to find the proteins that play a role in the development of this clinical pathogenesis and the effect of DM on protein expression. LF tissue retrieved from patients diagnosed with LSCS, some were also diagnosed with DM, were compared with LF from patients diagnosed with herniated nucleus pulposus (HNP). The tissues were analyzed by mass spectrometry for proteins profile alteration. We found that LF of LSCS/DM patients exhibited significantly higher levels of proteoglycan proteins and latent transforming growth factor β-binding protein (LTBP2 and LTBP4). Additionally, an increase of HTRA serine protease 1 and insulin-like growth factor binding protein-5 were noted. The higher fibrosis was also associated with proteins related to inflammation and slower tissue repair. Collagen 6 and transforming growth factor inhibitor are related to activation of the anti-inflammatory M2 pathway that is associated with tissue repair. The decrease of these proteins expression in LSCS/DM is associated with increased levels and activation of M1 pro-inflammatory pathways. Interestingly, C3 and C4b members of the complement complex and mannose receptor-like protein (CLEC18) paralogous proteins were detectable solely at the LSCS/DM patients' samples. Histology analysis shows that inflammatory was induced by the hyperglycemic conditions in diabetic patients involve in altering the matrix compositions. Thus, the protein profiles associated with inflammatory pathways affecting the LF suggested increasing susceptibility of developing the degeneration under hyperglycemic conditions.

Keywords:  diabetes mellitus; fibrosis; ligamentum flavum; mass spectrometry; spinal stenosis

DOI:  https://doi.org/10.1002/jcb.28451
mSystems. 2019 Jan-Feb;4(1):pii: e00337-18. [Epub ahead of print]4(1):

Evaluating Metagenomic Prediction of the Metaproteome in a 4.5-Year Study of a Patient with Crohn's Disease.

Robert H Mills, Yoshiki Vázquez-Baeza, Qiyun Zhu, Lingjing Jiang, James Gaffney, Greg Humphrey, Larry Smarr, Rob Knight, David J Gonzalez.

  Although genetic approaches are the standard in microbiome analysis, proteome-level information is largely absent. This discrepancy warrants a better understanding of the relationship between gene copy number and protein abundance, as this is crucial information for inferring protein-level changes from metagenomic data. As it remains unknown how metaproteomic systems evolve during dynamic disease states, we leveraged a 4.5-year fecal time series using samples from a single patient with colonic Crohn's disease. Utilizing multiplexed quantitative proteomics and shotgun metagenomic sequencing of eight time points in technical triplicate, we quantified over 29,000 protein groups and 110,000 genes and compared them to five protein biomarkers of disease activity. Broad-scale observations were consistent between data types, including overall clustering by principal-coordinate analysis and fluctuations in Gene Ontology terms related to Crohn's disease. Through linear regression, we determined genes and proteins fluctuating in conjunction with inflammatory metrics. We discovered conserved taxonomic differences relevant to Crohn's disease, including a negative association of Faecalibacterium and a positive association of Escherichia with calprotectin. Despite concordant associations of genera, the specific genes correlated with these metrics were drastically different between metagenomic and metaproteomic data sets. This resulted in the generation of unique functional interpretations dependent on the data type, with metaproteome evidence for previously investigated mechanisms of dysbiosis. An example of one such mechanism was a connection between urease enzymes, amino acid metabolism, and the local inflammation state within the patient. This proof-of-concept approach prompts further investigation of the metaproteome and its relationship with the metagenome in biologically complex systems such as the microbiome. IMPORTANCE A majority of current microbiome research relies heavily on DNA analysis. However, as the field moves toward understanding the microbial functions related to healthy and disease states, it is critical to evaluate how changes in DNA relate to changes in proteins, which are functional units of the genome. This study tracked the abundance of genes and proteins as they fluctuated during various inflammatory states in a 4.5-year study of a patient with colonic Crohn's disease. Our results indicate that despite a low level of correlation, taxonomic associations were consistent in the two data types. While there was overlap of the data types, several associations were uniquely discovered by analyzing the metaproteome component. This case study provides unique and important insights into the fundamental relationship between the genes and proteins of a single individual's fecal microbiome associated with clinical consequences.

Keywords:  colonic Crohn's disease; gut inflammation; inflammatory bowel disease; metagenomics; metaproteomics; microbiome; multiomics; proteomics; tandem mass tags; time series

DOI:  https://doi.org/10.1128/mSystems.00337-18
ACS Chem Neurosci. 2019 Feb 26.

Proteomics reveals profound metabolic changes in the alcohol use disorder brain.

Charmaine Enculescu, Edward D Kerr, K Y Benjamin Yeo, Gerhard Schenk, Marina R S Fortes, Benjamin L Schulz.

Changes in brain metabolism are a hallmark of Alcohol Use Disorder (AUD). Determining how AUD changes the brain proteome is critical for understanding the effects of alcohol consumption on biochemical processes in the brain. We used data-independent acquisition mass spectrometry proteomics to study differences in the abundance of proteins associated with AUD in pre-frontal lobe and motor cortex from autopsy brain. AUD had a substantial effect on the overall brain proteome exceeding the inherent differences between brain regions. Proteins associated with glycolysis, trafficking, the cytoskeleton, and excitotoxicity were altered in abundance in AUD. We observed extensive changes in the abundance of key metabolic enzymes, consistent with a switch from glucose to acetate utilization in the AUD brain. We propose that metabolic adaptations allowing efficient acetate utilization contribute to ethanol dependence in AUD.

DOI: https://doi.org/10.1021/acschemneuro.8b00660
Sci Rep. 2019 Feb 27. 9(1): 2905

Ibudilast sensitizes glioblastoma to temozolomide by targeting Macrophage Migration Inhibitory Factor (MIF).

Wendy Ha, Hatice Sevim-Nalkiran, Ashraf M Zaman, Kazuko Matsuda, Mustafa Khasraw, Anna K Nowak, Liping Chung, Robert C Baxter, Kerrie L McDonald.

Recurrence in patients with glioblastoma (GBM) is inevitable resulting in short survival times, even in patients with O-6-Methylguanine-DNA Methyltransferase (MGMT) methylation. Other pathways must be activated to escape from temozolomide (TMZ) treatment, however acquired resistance mechanisms to TMZ are not well understood. Herein, frozen tumors from 36 MGMT methylated patients grouped according to overall survival were extracted and proteins were profiled using surface-enhanced laser desorption/ionization (SELDI) with time-of flight (TOF) proteomics to identify low molecular weight proteins that associated with poor survival outcomes. Overexpression of macrophage migration inhibitory factor (MIF) was identified in human GBM specimens that were MGMT methylated but showed poor survival. This correlation was confirmed in an independent cohort of human GBM. MIF overexpression has been reported in several cancer types, including GBM. We repurposed ibudilast, a specific MIF inhibitor, and treated patient derived cell lines. Ibudilast showed modest anti-proliferative activity however, when combined with TMZ, significant synergism was observed, resulting in cell cycle arrest and apoptosis. In vivo, combined ibudilast and TMZ treatment of a patient derived xenograft (PDX) model resulted in significantly longer overall survival. Our findings have significant clinical implications for people with GBM. Since clinical trials involving ibudilast have shown no adverse side effects and the drug readily penetrates the blood brain barrier, treatment of GBM with this combination is clinically achievable.

DOI: https://doi.org/10.1038/s41598-019-39427-4
EBioMedicine. 2019 Feb 20. pii: S2352-3964(19)30090-8. [Epub ahead of print]

Re-assessment of the high-grade serous ovarian cancer phosphoproteome - Identification of kinase candidates in TCGA tumors for future clinical intervention.

Frank Rolfs, Sander R Piersma, Connie R Jimenez.

DOI: https://doi.org/10.1016/j.ebiom.2019.02.018
J Biol Regul Homeost Agents. 2019 Feb 25. 33(2):

AF1q gene polymorphism proteomic markers in herpes zoster-infected leukemia patients.

X Cheng, S L Zhao, H M Zhang.

Keywords: AF1q gene; gene polymorphism; iTraq; leukemia combined with herpes zoster; proteomic markers
FASEB J. 2019 Feb 27. fj201802090RR

Neutrophil exosomes enhance the skin autoinflammation in generalized pustular psoriasis via activating keratinocytes.

Shuai Shao, Hui Fang, Jingliang Zhang, Man Jiang, Ke Xue, Jingyi Ma, Jieyu Zhang, Jie Lei, Yangyang Zhang, Bing Li, Xu Yuan, Erle Dang, Gang Wang.

  Generalized pustular psoriasis (GPP) is a rare and severe inflammatory skin disease that can be life-threatening. Gene mutations are found in some cases, but its immune pathogenesis is largely unknown. Here, we observed that the neutrophil:lymphocyte ratio in patients with GPP was higher than that in healthy controls and decreased after effective treatment. Neutrophils isolated from patients with GPP induced higher expressions of inflammatory genes including IL-1β, IL-36G, IL-18, TNF-α, and C-X-C motif chemokine ligands in keratinocytes than normal neutrophils did. Moreover, neutrophils from patients with GPP secreted more exosomes than controls, which were then rapidly internalized by keratinocytes, increasing the expression of these inflammatory molecules via activating NF-κB and MAPK signaling pathways. The proteomic profiles in neutrophil exosomes further characterized functional proteins and identified olfactomedin 4 as the critical differentially expressed protein that mediates the autoimmune inflammatory responses of GPP. These results demonstrate that neutrophil exosomes have an immune-regulatory effect on keratinocytes, which modulates immune cell migration and autoinflammation in GPP.-Shao, S., Fang, H., Zhang, J., Jiang, M., Xue, K., Ma, J., Zhang, J., Lei, J., Zhang, Y., Li, B., Yuan, X., Dang, E., Wang, G. Neutrophil exosomes enhance the skin autoinflammation in generalized pustular psoriasis via activating keratinocytes.

Keywords:  GPP; OLFM4; autoimmune; neutrophils; vesicles

DOI:  https://doi.org/10.1096/fj.201802090RR